Long term potentiation (LTP) is a widely used model in studies on molecular and cellular mechanisms underlying synaptic plasticity. Elucidating the molecular and cellular properties underlying LTP is the unifying objective of this proposal. Previous studies have determined a postsynaptic site for LTP induction. However, the cellular residence for LTP expression, whether pre- and/or postsynaptic, remains heavily debated. A postsynaptic site of induction and a presynaptic site of expression have generated much interest in retrograde messengers that would communicate the induction of LTP from the post- to presynaptic neuron. Moreover, to preserve the synapse specificity of LTP many believe that there must be an activity dependent interaction of retrograde messengers with presynaptic terminals. We have begun to test these hypotheses by developing an induction protocol using glutamate iontophoresis that reliably induces LTP in hippocampal slices in the absence of presynaptic transmitter release, evoked stimulation and action potentials. Our results place constraints on the involvement of activity dependent presynaptic mechanisms and retrograde messengers in LTP induction and expression. Namely, these processes must function in the absence of many forms of presynaptic activity. LTP induced by glutamate iontophoresis ("ionto-LTP") will be used to elucidate pre- and postsynaptic mechanisms responsible for the induction and expression of LTP. We also propose experiments to test the involvement of retrograde messengers in ionto-LTP. LTP induction is dependent on Ca2+ activated protein kinases, however, the temporal duration of kinase activity required for LTP induction is unclear and controversial. Previous studies have simply assumed that the molecular mechanisms underlying LTP fall into two temporal stages, induction (seconds) and expression (hours). Our results indicate that there is critical window of protein kinase activity that occurs 2-10 min after the induction event. We believe this period of protein kinase activity is a stabilization or consolidation stage that occurs after induction, but is required for LTP expression. We plan to determine the temporal requirement for protein kinase activity during LTP consolidation, and to identify which specific protein kinase(s) is/are important during this period. The elucidation of the temporal requirement for kinase dependent processes will be instrumental to future studies on molecular changes responsible for LTP expression. Our preliminary results also indicate a role for protein kinases in regulating short-term synaptic plasticity and experiments are proposed to test kinase involvement in paired-pulse facilitation and post-tetanic potentiation. These studies are a multidisciplinary approach to elucidating mechanisms in plasticity and should provide new and important information about the role of second messenger-dependent events responsible for LTP expression.